鱼类病原菌迟钝爱德华氏菌功能RNA组研究

Edwardsiella tarda is the etiological agent of edwardsiellosis, characterized by systemic hemorrhagic septicemia and internal abscesses, leading to mass mortality outbreaks in many freshwater and marine fish. It has caused devastating economic losses in USA, Europe and Asia. Systematically exploring niche adaptation, virulence related genes and the interaction networks of these genes will help us to understand pathogenic mechanisms and niche adaptation systems of this pathogenic microorganism. In order to uncover the mechanisms of virulence regulation and niche adaptation regulation of this pathogen, we plan to systematic investigate the transcriptomes of different E. tarda strains isolated from various hosts and at various physiological states. Following detecting and determining the expression profiles of mRNA and non-coding RNA (ncRNA) in different states, we hope to clarify the roles of the critical mRNA/ncRNAs of the pathogen in its invasion and survival in fish and survival at various wild environments. We will also investigate the regulatory roles of critical mRNA/ncRNAs with molecular genetic methods such as site-directed deletion and overexpression to test whether these candidate genes influence on virulence and adaptation processes in E. tarda and finally establish the virulence regulation networks and niche adaptation response networks based on protein-protein and protein-functional RNA interactions.

迟钝爱德华氏菌是我国北方沿海地区养殖鲆鲽等经济鱼种主要的病原菌之一，能引起鱼体肠道出血性败血症并造成大量死亡，已经对美国，欧洲及亚洲等国家造成大量经济损失。对其环境适应性及致病相关基因及其互作方式进行研究，成为防控水产养殖病原菌的重要基础。本项目拟系统研究不同来源迟钝爱德华氏菌从体外环境到鱼类宿主环境并造成鱼类疾病暴发过程中各生理状态下RNA组的变化规律，用于研究该菌在不同条件下精细环境响应网络，以及不同菌株不同的毒力蛋白调控模式。通过分析不同状态下mRNA及非编码RNA的表达谱，在分子水平上鉴定迟钝爱德华氏菌侵染鱼体和在鱼体内存活中起关键调控作用的功能RNA分子，通过基因缺失及过表达等方法分析功能RNA的生物学含义并揭示迟钝爱德华氏菌致病性的机理，并构建迟钝爱德华氏菌中由mRNA，非编码RNA和蛋白质构成的环境适应/毒力调控网络。

迟钝爱德华氏菌也称为杀鱼爱德华氏菌(Edwardsiella piscicida)，是一种广宿主的革兰氏阴性致病菌，能造成许多重要经济养殖鱼种的出血性败血症，更好地了解其致病机制将有助于开发安全高效的疫苗来防控爱德华氏菌病。目前已经鉴定出多个毒力因子参与到E. piscicida的致病过程中，其中III型分泌系统(T3SS)协助其逃避宿主吞噬细胞的杀伤并在胞内增殖，VI型分泌系统(T6SS)也协助其在宿主体内定植。EsrA-EsrB双组份系统对T3SS和T6SS有重要的调控作用，但EsrB在E. piscicida中对毒力以及生理的全局调控作用尚不清楚。本课题对比了E. piscicida野生株和ΔesrB缺失株在不同条件下的转录谱，分析了差异表达基因，寻找受EsrB调控的宿主适应性及毒力相关基因。我们鉴定了6个新的效应蛋白，它们通过T3SS转运至宿主细胞中，与病原菌的定植与毒力相关。在此基础上，我们鉴定了2个受EsrB直接调控的基因，在它们的启动子区域发现了结合EsrB的区域(Binding box)。总之，本课题较好地完成了项目设定的研究目标，阐明了杀鱼爱德华氏菌中关键毒力调控蛋白EsrB对毒力基因、代谢途径以及应激适应相关基因的全局调控作用，也阐明了该菌在模拟体内和体外环境中的RNA组的变化，为理解该菌的毒力调控机制、感染过程以及新型疫苗的开发打下了良好基础。受该项目资助，本课题发表SCI学术论文两篇，其中在Virulence和FEMS Microbiology Letters各一篇。